Abstract Objective In patients with suspected coronary artery disease (CAD) and intermediate pretest probability non-invasive stress-testing is recommended in current guidelines. However, sensitivity and specificity of these tests are limited. We aimed to investigate the additional diagnostic value of high-sensitivity troponin I concentrations measured before and after stress-testing. Methods We included patients undergoing non-invasive stress-testing in a prospective cohort study. All patients provided written informed consent, were at least 18 years old and underwent either stress-echocardiography, stress-MRI or myocardial perfusion scintigraphy. Baseline parameters were collected by questionnaire and chart review. Blood samples were collected before, and one hour after the stress-test. Troponin was measured using the Abbott Architect high-sensitivity troponin I (hs-TnI) assay. The diagnostic performance to predict a pathological stress-test was evaluated by calculation of the area under the ROC curve (AUC) using baseline hs-TnI concentrations and the absolute change after one hour. An optimal cutoff was determined by maximization of the Youden Index. Univariate odds ratios (OR) were calculated to identify predictors for a pathological stress-test. Additionally ORs for the optimal hs-TnI cutoff were calculated in a multivarite regression with adjustment for baseline variables. Results In total 391 patients with a median age of 70 years were included. 66.8% were males, 83.4% had prevalent hypertension, 55.9% had dyslipidemia and 55.4% had a prior history of CAD. A pathological stress-test was reported in 21.5% and these patients had a higher cardiovascular risk profile, compared to patients with a non-pathological stress-test. The baseline hs-TnI concentrations were 4.7 ng/L in the overall population, 4.2 ng/L in patients with a non-pathological stress-test and 8.4 ng/L (p<0.001) in patients with a pathological stress-test. There was no significant hs-TnI changes one hour after stress-testing. The AUC for the baseline hs-TnI was 0.65 and the optimal cutoff was determined at 7.7 ng/L. Most cardiovascular risk factors were predictors for a pathological stress-test. The OR for an elevated hs-TnI above the optimized cutoff was 3.34 (95% CI 2.03, 5.52). In a multivariate model a baseline hs-TnI above 7.7 ng/L showed an OR of 2.07 (95% CI 1.04, 4.12; p-value 0.039) Conclusion In patients undergoing non-invasive stress-testing, hs-TnI concentrations did not significantly change one hour after exercise. However, an elevated troponin concentration is an independent predictor for a pathological stress-test.